细粒棘球绦虫抗原EgAgB8／1和EgAgB8／3亚单位基因在虫体发育过程中的阶段性表达  

Echinococcus granulosus:developmental stage-specific expression of antigen B 8-ku-subunits 1 and 3

作　　者：张海涛[1] 马秀敏[1,2] 吾拉木·马木提[1,2] 马海梅[1,2] 贾海英[1] 曹春宝[1] 丁剑冰[1,2] 温浩[1] 

机构地区：[1]新疆医科大学第一附属医院,新疆包虫病VIP实验室,新疆乌鲁木齐830011 [2]新疆医科大学基础医学院

出　　处：《中国病原生物学杂志》2009年第5期355-358,共4页Journal of Pathogen Biology

基　　金：国家自然科学基金资助项目(No.30760229);科技部“863计划”项目(No.2007AA02Z411);新疆自治区重点实验室开放课题(No.XJDX0202-2007-01)

摘　　要：目的研究细粒棘球绦虫抗原B(EgAgB)8 ku 1和3亚单位基因家族成员(EgAgB8/1、EgAgB8/3)在虫体发育各阶段(虫卵、棘球蚴生发层、原头蚴及成虫)基因表达的差异,为包虫感染中间宿主和终末宿主的免疫学诊断筛选期特异性优势表达抗原。方法分别从细粒棘球绦虫成虫、虫卵、原头蚴和棘球蚴生发层提取总RNA,用Avian Myelo-blastosis Virus Reverse Transcriptase XL反转录成cDNA,根据EgAgB8/1和EgAgB8/3序列设计跨越内含子的基因特异性引物。基因表达差异应用SYBR GreenⅠreal-time PCR进行定量分析,并用内参对照基因actinⅡ来标准化定量结果。结果Real-time PCR检测显示,EgAgB8/1在棘球蚴生发层大量表达,SQ/Actin=68.874;EgAgB8/3在成虫阶段大量表达,SQ/Actin=1 905.512。结论EgAgB 2个亚单位基因的表达在虫体发育各个阶段显现出明显差异。提示EgAgB8/1可作为包虫感染中间宿主免疫学诊断最佳候选目的抗原,而EgAgB8/3可作为包虫感染终末宿主免疫学诊断的靶抗原。EgAgB 2个亚单位基因的阶段差异表达可能与其虫体发育不同阶段特有的生物学功能有关。Objective To understand the expression profile of 8-ku 1 and 3 of antigen B （EgAgB8/1, EgAgB8/3） at different developmental stages （eggs, hydatid cyst germinal layer, protoscoleces and adult worms） of Echinococcus granulosus and to select the predominant target antigen for the immunodiagnosis of echinococcal infection in intermediate hosts and definitive hosts. Methods Total RNA was isolated from fresh samples of E. granulosus adult worms, eggs, proto-scoleces and hydatid cyst germinal layers. First stand cDNA was synthesized using Avian Myeloblastosis Virus Reverse Transeriptase XL. The expression level of the target genes at different developmental stages of the parasite was quantified by SYBR Green Ⅰ real-time PCR with sequence-specific intron-spanning primers designed according to gene sequences of EgAgB8/1 and EgAgBS/3. Relative gene expression results were normalized by parallel studies with internal control gene actin Ⅱ. Results Real-time PCR results showed that the gene expression level of EgAgB8/1 were up regulated in the hydatid cyst germinal layer （SQ/Actin= 68. 874） compared with that in eggs （SQ/Actin= 2. 089）, protoscoleces （SQ/ Actin=0. 56） and adult worms （SQ/Actin= 0. 007）. EgAgB8/3 were over expressed in adult worms （SQ/Actin= 1 905. 512） relative to other developmental stages. Conclusion Two subunits of EgAgB seemed to be differentially expressed at different developmental stages of the parasite. According to our data, EgAgB8/1 may be the best choice of target antigen for serodiagnosis of echinococcal infection in intermediate hosts, including humans, while EgAgB8/3 is the best choice in definitive hosts. The stage-specific expression patterns of these two genes may be related to different functions of this antigen during the parasite life cycle; this should be further evaluated in subsequent studies.

关 键 词：细粒棘球绦虫 抗原B亚单位 荧光定量PCR 表达 

分 类 号：R383.33[医药卫生—医学寄生虫学]